PROJECT SUMMARY Filoviruses that circulate in the Eastern Hemisphere and several arenaviruses that circulate in the Western hemisphere cause human hemorrhagic fevers with case fatality rates (15% to 90%). Most lack treatment options. The research project proposed here is an effort to use the blood of survivors of viral hemorrhagic fevers to understand the basic molecular features, on both the human host and pathogen side, that make transfusion of antibodies (passive immunity) more or less effective in treating human viral hemorrhagic fevers. To achieve this goal, we propose to compare the human antibody response to filoviruses and arenaviruses. We chose to compare both groups of viruses because for Junn virus infection, transfusion of survivor plasma is highly effective and routinely used clinically ? it decreases the case fatality rate of infection to less than 1%. In contrast, while passive immunity has also shown promise in animal models against Ebola virus, particularly with the use of a three antibody-cocktail called ZMapp, it has yet to be clearly demonstrated as effective in humans. We have recruited survivors of Junn virus, Ebola virus, and Marburg virus infections for blood donation. In Aim 1, we will recover the genes encoding antibodies that react against the arenavirus and filovirus surface glycoprotein (the target of antibodies) from the blood of these survivors. With the sequences of antibody genes, we will produce large quantities of each identified antibody as recombinant proteins and study these in binding and viral neutralization assays. In Aim 2, we will determine antibody-glycoprotein structure using X-ray crystallography and cryo- electron (Cryo-EM) microscopy. In Aim 3, we will study the antibodies' non-neutralizing antiviral activity in cell-based assays and in small animal models working closely with our collaborators. We will also determine if we can alleviate a known side effect of passive immunity against Junn virus ? development, in 10% of treated survivors, of a late neurological syndrome that probably occurs because of viral replication in their central nervous system. The basic findings from our research will help us design the next generation of passive immunity strategies against emerging viruses that cause severe human diseases ? that is, specifically tailored monoclonal antibody therapies in which purified antibodies of known activity are included.